2009
DOI: 10.1111/j.1365-3083.2009.02338.x
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Structural Difference in the Complement Activation Site of Human IgG1 and IgG3

Abstract: The C1q binding epicentre on IgG molecules involves residues Asp270, Lys322, Pro329 and Pro331 in the CH2 domain. IgG1 and IgG3 are usually the most efficient of the four human IgG subclasses in activating complement and they both share all these residues. To reveal possible differences in the structural requirement for complement activation, we created a number of NIP (5‐iodo‐4‐hydroxy‐3‐nitro‐phenacetyl) specific IgG1 and IgG3 antibodies with parallel mutations in or near the putative C1q binding site. The m… Show more

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Cited by 41 publications
(43 citation statements)
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“…Human IgG3 has an extended hinge region (63 amino acids, as opposed to 15 amino acids for IgG1) (26), which might allow for greater flexibility of the Fc portion of the IgG3 molecule and more efficient engagement of C1q when fHbp is sparsely exposed. This hypothesis, however, is not supported by studies of mutant human IgG3 molecules with truncated hinge regions where the greater ability of IgG3 than IgG1 to activate complement-mediated lysis of red cells with low hapten density was independent of the hinge region (29). Based upon data from mutant IgG1 and IgG3 molecules with substitutions of single amino acid residues contributing to C1q binding (29), subtle structural differences in the CH2 domain appear to be responsible for the greater ability of IgG3 to engage C1q and activate the classical complement pathway when the target antigen is sparse.…”
Section: Discussionmentioning
confidence: 63%
“…Human IgG3 has an extended hinge region (63 amino acids, as opposed to 15 amino acids for IgG1) (26), which might allow for greater flexibility of the Fc portion of the IgG3 molecule and more efficient engagement of C1q when fHbp is sparsely exposed. This hypothesis, however, is not supported by studies of mutant human IgG3 molecules with truncated hinge regions where the greater ability of IgG3 than IgG1 to activate complement-mediated lysis of red cells with low hapten density was independent of the hinge region (29). Based upon data from mutant IgG1 and IgG3 molecules with substitutions of single amino acid residues contributing to C1q binding (29), subtle structural differences in the CH2 domain appear to be responsible for the greater ability of IgG3 to engage C1q and activate the classical complement pathway when the target antigen is sparse.…”
Section: Discussionmentioning
confidence: 63%
“…However, IgG1 is much more dependent on Asp270 than IgG3, and thus, the IgG3D270A mutant (which carries a mutation from aspartic acid to alanine at position 270) maintains most of the complement activation activity, while the IgG1D270A mutant is devoid of complement activation activity, and this difference is independent of the long hinge region of IgG3 (57).…”
Section: Discussionmentioning
confidence: 99%
“…Human IgG1 and IgG3 share the amino acid residues that make up the epicenter of the complement activation site on the Fc part of the molecules (Asp270, Lys322, Pro329, and Pro331), but parallel mutations in other common surrounding and shared amino acids in IgG1 and IgG3 indicate that there are subtle differences in the complement activation site of human IgG1 and IgG3 (57). However, IgG1 is much more dependent on Asp270 than IgG3, and thus, the IgG3D270A mutant (which carries a mutation from aspartic acid to alanine at position 270) maintains most of the complement activation activity, while the IgG1D270A mutant is devoid of complement activation activity, and this difference is independent of the long hinge region of IgG3 (57).…”
Section: Discussionmentioning
confidence: 99%
“…7D, 7E). As controls, we included hIgG1 with a single point mutation (P329A) within the C H 2 domain at the core of the C1q binding site (47), which completely eliminated hC1q binding, as well as hIgG1 with mutation of either of two leucine residues in the lower hinge (L234A and L235A), crucial for FcgR binding (54), which moderately reduced binding (Supplemental Fig. 3E-H).…”
Section: Binding Of the Fc-engineered Higg1 Variants To Hc1qmentioning
confidence: 99%
“…The vector contains the constant H chain gene from hIgG1 with specificity for the hapten 5-iodo-4-hydroxy-3-nitro-phenacetyl (NIP), and it was used as a template for subcloning of DNA fragments encoding (synthesized by GenScript) a panel of Fc mutants using the restriction sites AgeI and SfiI (C H 2 mutations) or SfiI and BamHI (C H 3 mutations): M252Y/ S254T/T256E (MST), M428L/N434S (MN), H433K/N434F (HN), and M252Y/S254T/T256E/H433K/N434F (MST/HN). Vectors encoding the constant H chain gene from hIgG1 with the mutations P329A, L234A, and L235A have previously been described (25,47). The Abs were produced using HEK293E cells by transient cotransfection of the vectors together with a vector encoding the mouse l L chain with NIP specificity (pLNOH2-NIPlLC-oriP) using Lipofectamine 2000 (Invitrogen) following the manufacturer's instructions.…”
Section: Cell Culturementioning
confidence: 99%